IL-15 that activates natural killer (NK) cells, has potential applications in cancer immunotherapy. Based on rhesus macaques' experiments, we performed a first-in-human phase I dose-escalation trial of Escherichia coli (E. coli) produced rhIL-15 with NCI as the sponsor, Dr. Kevin Conlon as the Principal Investigator (PI) and Dr. Thomas Waldmann as Study Chairman and IND holder. This study was completed with 5 patients having finished their course of therapy at 3.0 mcg/kg/day, 4 patients having entered at 1.0 mcg/kg/day and 5 patients at the 0.3 mcg/kg/day dose of 12 daily infusions. Following the bolus i.v. infusion of IL-15 the serum concentrations of IL-15 at 10 minutes following infusion ranged from 20,000 to 90,000 picograms/mL levels-- sufficient to signal through the IL-2/IL-15R beta and common gamma receptors shared with IL-2. The pharmacokinetics for various parameters for the 3.0, 1.0 and 0.3 mcg/kg/day doses resulted in a Cmax of 47,800 +/- 18,300 picograms/mL, 15,900 +/- 19,000 picograms/mL and 12,060 +/- 350 pg/mL respectively. The half-lives (T 1/2 ) were very similar for the three dose levels 2.4 +/- 0.5 hours, 2.7 +/- 0.13 hours and 2.7 +/- 0.67 hours. With a bolus infusion of 3.0 mcg/kg/day there was a pattern of fever beginning 2 to 2.5 hours following the start of the rhIL-15 infusions, peaking reliably at the 3-hour timepoint. Rigors occurred at the 4-hour timepoint. Rigors and fever were also observed at the 1.0 mcg/kg/day dose but were less severe. Up to 50-fold increases of serum levels of inflammatory cytokines in particular IL-6, IL-8 and IFN gamma were observed. There was minimal fever observed at the 0.3 mcg/day dose. These toxicities were associated with maximum elevations at 4 to 8 hours post-bolus infusion of IL-15 in the serum concentrations of inflammatory cytokines (e.g. IL-6, Il-1, IL-8, IL-10 and IFN-gamma). Two patients manifested dose-limiting toxicity (DLT) at both the 3.0 mcg/kg/day (hypotension) and at the 1.0 mcg/kg/day (abnormalities of liver function). However the 0.3 mcg/kg/day dose has proven to be without DLT in the 5 patients examined. Flow cytometry of peripheral blood lymphocytes revealed a dramatic efflux of NK and memory CD8 T cells from the circulating blood within minutes upon IL-15 administration, followed by influx and hyperproliferation yielding 10-fold expansions of NK and gamma delta T cells that ultimately returned to baseline. The pharmacokinetics of IL-15 following bolus infusion, discussed above, are clearly not optimal with exceedingly high levels initially that may have caused the toxicity. Therefore a trial has been initiated using continuous intravenous infusion of rhIL-15. With the Cancer Immunotherapy Network (CITN) a trial has been initiated in a dose-escalation trial for a 10-day period. Furthermore, a subcutaneous dosing strategy for IL-15 has been initiated as well. A second limitation in the present trial of rhIL-15 is that the levels of IL-15R alpha required for optimal IL-15 action observed in the patients prior to and during the IL-15 infusions were exceptionally low. Therefore we propose to translate our preclinical studies in murine tumor models by initiating a combination agent clinical trial that involves the use of an agonistic antibody to CD40 to induce IL-15R alpha expression in conjunction with IL-15 administration. Furthermore, our present and future plans include administration of Il-15 in conjunction with anti-PD1 with anti-CTLA-4 monoclonal antibodies to remove these checkpoints on the immune response. Collectively the 2-decade long scientific odyssey that involved the discovery and development of IL-15 by the Waldmann Group has accelerated progress in the field of Immunobiology that focuses on the role of the common gamma cytokines in the normal regulation of NK and CD8 T-cell homeostasis and in disorders in this regulation in disease states. Furthermore the completed initial trial evaluating IL-15 in patients with metastatic malignancy has provided the phase I goal information required for the initiation of a broad range of studies using IL-15 in clinical trials by intramural and extramural NCI supported groups such as the Cancer Immunotherapy Network (CITN).